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. Author manuscript; available in PMC: 2009 Jun 8.
Published in final edited form as: Biochem J. 2008 May 1;411(3):523–530. doi: 10.1042/BJ20071436

Figure 4. In vitro dsDNA cleavage assay.

Figure 4

A) Schematic diagram of the 3'-labeled 33-mer dsDNA substrate (32P-dideoxyadenosine label is indicated by *), with Top1 (black circle) making a covalent bond to the DNA backbone resulting in the 23-mer oligonucleotide cleavage product. B) Top1 dsDNA cleavage assay with 10μM CPT and 10 nM dsDNA. Reaction products were separated in denaturing PAGE. Lane C, control reaction with no added Top1; lane 1, Top69NLS (7.5 nM); lane 2, Top68 (7.5 nM); lane 3, Top68W/A (30 nM). The 33-mer oligonucleotide is indicated on the left, as is the 23-mer oligonucleotide cleavage product.